PTEN Activation by DNA Damage Induces Protective Autophagy in Response to Cucurbitacin B in Hepatocellular Carcinoma Cells.

Yanan Niu,Xiuping Chen,Lixia Pei,Jin-Jian Lu,Dik-Lung Ma,Chung-Hang Leung,Wen Sun

doi:10.1155/2016/4313204

Abstract

Cucurbitacin B (Cuc B), a natural product, induced both protective autophagy and DNA damage mediated by ROS while the detailed mechanisms remain unclear. This study explored the mechanism of Cuc B-induced DNA damage and autophagy. Cuc B decreased cell viability in concentration- and time-dependent manners. Cuc B caused long comet tails and increased expression of γ-H2AX, phosphorylation of ATM/ATR, and Chk1/Chk2. Cuc B induced autophagy as evidenced by monodansylcadaverine (MDC) staining, increased expression of LC3II, phosphorylated ULK1, and decreased expression of phosphorylated AKT, mTOR. Cuc B induced apoptosis mediated by Bcl-2 family proteins and caspase activation. Furthermore, Cuc B induced ROS formation, which was inhibited by N-acetyl-L-cysteine (NAC). NAC pretreatment dramatically reversed Cuc B-induced DNA damage, autophagy, and apoptosis. Cuc B-induced apoptosis was reversed by NAC but enhanced by 3-methyladenine (3-MA), chloroquine (CQ), and silencing phosphatase and tensin homolog (PTEN). 3-MA and CQ showed no effect on Cuc B-induced DNA damage. In addition, Cuc B increased PTEN phosphorylation and silence PTEN restored Cuc B-induced autophagic protein expressions without affecting DNA damage. In summary, Cuc B induced DNA damage, apoptosis, and protective autophagy mediated by ROS. PTEN activation in response to DNA damage bridged DNA damage and prosurvival autophagy.

Highlights

Programmed cell death (PCD), a process carried out in a regulated manner, ubiquitously occurs throughout most multicellular organisms’ lifespan
Accumulated evidence showed that though apoptosis and autophagy were executed through distinct signaling pathways, overlapping signals were engaged in response to specific stimuli [1]
We firstly reported that Cuc B induced DNA damage mediated by Reactive oxygen species (ROS) in A549, K562, and MCF-7 cells [23, 27, 28]

Summary

Introduction

Programmed cell death (PCD), a process carried out in a regulated manner, ubiquitously occurs throughout most multicellular organisms’ lifespan. Accumulated evidence showed that though apoptosis and autophagy were executed through distinct signaling pathways, overlapping signals were engaged in response to specific stimuli [1]. This crosstalk could be mediated by the interactions between Beclin-1 and Bcl-2/Bcl-xL and between FADD and Atg, caspase- and calpain-mediated cleavage of autophagy-related proteins, and autophagic degradation of caspases [9,10,11,12,13]. We firstly reported that Cuc B induced DNA damage mediated by ROS in A549, K562, and MCF-7 cells [23, 27, 28]. The DNA damage activated phosphatase and tensin homolog (PTEN) bridged DNA damage and autophagy

Materials and Methods

Results

Discussion